Diversity of aster yellows phytoplasmas in lettuce [electronic resource] /
Abstract: Seven aster yellows phytoplasma (AYP) strains were isolated from lettuce in Ohio and characterized based on host symptoms and by polymerase chain reaction (PCR) and restriction fragment length polymorphism analysis of PCR products (PCR-RFLP). AY-WB is the only strain that belongs to the 16SrI-A subgroup whereas the others belong to the 16SrI-B subgroup. AY-WB can be distinguished by PCR with strain specific primers BF/BR. AY-WB, AY-S, AY-BW and AY-SS can be distinguished from one another either by PCR-RFLP of primers AY19p/AY19m-amplified products or by multiplex PCR. Primer typing can be used to differentiate most strains except AY-BW and AY-SG. Phylogenetic analysis of partial sequences of 16S rRNA gene and 16S-23S spacer regions showed that all the strains were in the same cluster with most AYPs from the Genbank database. AYP was detected by PCR at the growing points of lettuce plants seven days before symptom appearance, indicating that aster yellows symptoms are closely related to the presence of phytoplasmas. AY-S spread faster and more widely than AY-BD2 within lettuce plants. A shotgun library was constructed by Integrated Genomics Inc. with AY-WB genomic DNA isolated using pulse field gel electrophoresis. The 800 kb genome was sequenced with 8.6-fold coverage at sequence saturation. Sixteen contigs have been constructed that cover 87.5% of the genome with 26.9% GC. Sixty-seven percent of the genome consists of coding sequences. Eleven transposase-coding regions were identified in length of 237?963 bp with GC content 23.4?27.4%. Seven of the transposase-coding regions contain two-linked ORFs, AYP167-166, AYP349-348, AYP503-504, AYP266-265, AYP706-705, AYP207-206 and AYP316-315, most overhung by 11-27 bp imperfect inverted repeats (IR). Most of the two-linked ORFs are out-of-phase coding regions, suggesting a frameshifting mechanism involved in translation process and four of them generated direct repeats (DR). AYP763, AYP199 and AYP015 are single ORF coding regions and all lack IRs. AYP231-2-3 contains three ORFs with a 39 bp deletion. PCR with primers based on sequences flanking transposase-coding regions generated the DNA fragments only from DNA extracts of aster yellows infected lettuce and leafhoppers not of healthy ones.
School:The Ohio State University
School Location:USA - Ohio
Source Type:Master's Thesis
Keywords:lettuce aster yellows plant viruses
Date of Publication: